The long term objective of this proposal is to understand the etiology and pathogenesis of Alzheimer disease (AD). Increasing evidence suggests that in AD the phosphorylation state of several neuronal proteins are altered. Such abnormal phosphorylation apparently lead to a disruption of different cellular functions and hence eventual death of neurons. In particular the microtubule-associated protein tau has been shown to be present in a hyperphosphorylated state in the paired helical filaments (PHF) of AD brain. Recent evidence suggests that the hyperphosphorylation of PHF tau may be catalyzed largely. but not exclusively, by proline-directed protein kinases (PDPKs). There are three well known PDPKs: MAP kinase, cdc2 kinase, and glycogen synthase kinase 3. The identities of the PDPKs and non-PDPKs responsible for the in vivo hyperphosphorylation of PHF tau are presently unclear. The specific aims of this proposal are: 1) To compare the phosphorylation of tau by the three PDPKs. The kinetics of phosphorylation and the sites phosphorylated on tau will be compared by peptide mapping and by different phosphorylation site-specific anti-tau antibodies; 2) To investigate the role of non-PDPKs in tau hyperphosphorylation. Six non- PDPKs will be used to prepare six species of phosphorylated tau. The further phosphorylation of these tau species by the PDPKs will be compared; 3) To use different synthetic peptides containing known phosphorylation sites of tau to establish the identities of the protein kinases responsible for their phosphorylation; and 4) To measure the levels of the PDPKs in both the cytosol and particulate fractions of control and AD brains. Both kinase activity and kinase protein levels will be measured. These studies should help clarify the identities of the kinase(s) responsible for the hyperphosphorylation of PHF tau.